The present invention relates to methods of preventing or reducing in a subject rejection of a non-syngeneic graft which comprise administering to the subject “third-party” tolerogenic cells which are non-syngeneic with both the recipient and the graft. The present invention further relates to articles of manufacture which comprise such tolerogenic cells which can be used for practicing such methods. More particularly, the present invention relates to methods of preventing or reducing in a subject rejection of an allogeneic graft which comprise administering to the subject CD4+CD25+ cells which may be allogeneic and fully MHC-mismatched with both the subject and the graft. The present invention further particularly relates to methods of preventing or reducing in a subject rejection of a xenogeneic graft which comprise administering to the subject CD4+CD25+ cells which are allogeneic and fully MHC-mismatched with the subject. By virtue of enabling for the first time use of such tolerogenic cells for preventing or reducing non-syngeneic graft rejection the present invention provides a routinely applicable and convenient method of treating a disease by allogeneic or xenogeneic graft administration/transplantation without or with reduced concomitant graft rejection.
Diseases which are treatable using treatment regimens involving administration of non-syngeneic grafts, such as bone marrow grafts, or pancreatic grafts, comprise numerous diseases which are characterized by significant mortality and morbidity, and for which no satisfactory/optimal treatments are presently available.
Examples of diseases which are treatable via treatment regimens involving administration of bone marrow cell allografts include hematopoietic malignancies whose treatment may be performed via radiotherapy/chemotherapy resulting in myeloablation/myeloreduction followed by bone marrow/hematopoietic stem cell administration for hematopoietic reconstitution. Such diseases also include those, such as organ failure, which are treatable by administration of non-syngeneic donor-derived organs, since engraftment of donor-derived grafts in general can be facilitated by establishment of hematopoietic chimerism in the graft recipient by adjunct donor-derived bone marrow transplantation.
Transplantation of pancreatic grafts is a potentially optimal method of treating pancreatic diseases such as diabetes. Diabetes is a debilitating and potentially lethal disease that develops in nearly 5 percent of the world's population. In the United States alone, an estimated 18 million people have diabetes mellitus, and each year about 1 million Americans aged 20 or older are diagnosed with the disease. It is the sixth leading cause of death in the US and is responsible for over 200,000 deaths a year. People with diabetes have a shortage of insulin or a reduced ability to use insulin, the hormone regulates blood glucose levels. In mammals the pancreas is responsible for the production and secretion of insulin. The standard therapy for diabetes, daily injections of insulin, does not satisfactorily prevent the debilitating and lethal consequences of this disease.
Significant obstacles to practicing therapeutic administration of allografts, the standard type of therapeutic graft employed, include the unavailability of suitably immunologically haplotype-matched grafts, and common complications, such as graft rejection, graft-versus-host disease (GVHD), and toxicity of immunosuppressant drugs such as cyclosporin A.
An alternative to allograft transplantation involves xenograft transplantation, i.e., transplantation of animal-derived grafts, in particular porcine grafts, which are considered a potentially optimal animal alternative to human grafts. The great advantages of using xenografts for transplantation are their availability on demand to all patients in need of transplantation, as well as avoidance of the medical and ethical burden of harvesting grafts from live or cadaveric human donors. However, to date, xenogeneic grafts have been ruled out for human transplantation due to their heretofore insurmountable immunological incompatibility with human recipients.
Bone marrow transplantation following supralethal radiochemotherapy is associated with dangerous infections due to the relatively slow rate of immune reconstitution during the first year after transplantation (Davison, G. M. et al., 2000. Transplantation 69:1341; Mencacci, A. et al., 2001. Blood 97:1483; Pan, L. et al., 1995. Blood 86:4422; Small, T. N. et al., 1999. Blood 93:467; Volpi, I. et al., 2001. Blood 97:2514; Weinberg, K. et al., 2001. Blood 97:1458). Thus, treatment methods involving reduced-intensity conditioning, associated with less severe immune ablation, are highly desirable, for example, for the treatment of a variety of nonmalignant diseases or for the induction of “mixed chimerism” as a prelude for cell therapy in cancer or in organ transplantation. However, the marked level of host hematopoietic and immune cells surviving mild preparatory regimens represents a difficult barrier for the engraftment of donor cells. In patients with advanced hematologic malignancies who cannot withstand myeloablative conditioning because of age and/or performance status, attempts have been made to develop low toxicity conditioning protocols in conjunction with human leukocyte antigen (HLA)-matched transplants (Giralt, S. et al., 1997. Blood 89:4531; Slavin, S. et al., 1998. Blood 91:756; McSweeney, P. A. et al., 2001. Blood 97:3390; Giralt, S. et al., 2001. Blood 97:631). Potent posttransplantation immunosuppression and the presence of large numbers of alloreactive T-cells in the graft enabled a high rate of engraftment. However, graft-versus-host disease (GVHD), particularly lethal chronic GVHD, remains a major obstacle (McSweeney, P. A. et al., 2001. Blood 97:3390; Einsele, H. et al., 2003. Br J Haematol 121:411; Bishop, M. R. et al., 2003. Biol Blood Marrow Transplant 9:162; Levine, J. E. et al., 2003. Biol Blood Marrow Transplant 9:189). While in high-risk leukemia such transplant-related mortality is acceptable, it would be totally intolerable if applied to patients with long life expectancy. Thus, the use of purified allogeneic stem cells, that do not pose any risk for GVHD and can continuously present donor-type antigens in the host thymus, thereby inducing durable tolerance to donor cells or tissues, represents one of the most desirable goals in transplantation biology. One approach to overcoming immune rejection of incompatible stem cells rigorously depleted of T-cells made use initially of increased doses of T-cell-depleted bone marrow (Aversa, F. et al., 1994. Blood 84:3948; Lapidot, T. et al., 1989. Blood 73:2025; Bachar-Lustig, E. et al., 1995. Nature Medicine 1:1268; Reisner, Y., and M. F. Martelli., 1995. Immunol Today 16:437) and rats (Uharek, L. et al., 1992. Blood 79:1612). Subsequently the cell-dose escalation concept was also shown with purified stem cells (Uchida, N. et al., 1998. J. Clin. Invest. 101:961; Aversa, F. et al., 1998. New Eng. J. Med 339:1186; Bachar-Lustig, E. et al., 1999. Blood 94:3212; Reisner, Y., and M. F. Martelli, 1999. Immunol Today 20:343). However, although this modality has become a clinical reality in the treatment of patients with leukemia conditioned by intensive chemotherapy, it has been suggested in studies in mice (Bachar-Lustig, E. et al., 1999. Blood 94:3212) and nonhuman primates (X. Yao, unpublished data, July 2001) that the number of hematopoietic precursors required to overcome the immune barrier in hosts pretreated with sublethal regimens cannot be attained with the state-of-the-art technology for stem cell mobilization. It has been demonstrated that when purified CD34+ cells are added to bulk mixed-lymphocyte reactions these cells suppress cytotoxic T-lymphocytes (CTLs) against matched stimulators but not against stimulators from a third-party (Rachamim, N. et al., 1998. Transplantation 65:1386). These results, which were further confirmed and extended by Gur et al (Gur, H. et al., 2002. Blood 99:4174; Gur, H. et al., 2005. Blood 105:2585) strongly indicated that cells within the human CD34+ population are endowed with potent veto activity. Considering that the number of human CD34+ cells that can be harvested is limited, the availability of other types of veto cells or immunoregulatory cells is crucial for further application of allogeneic stem cell transplantation under reduced intensity conditioning.
A potentially optimal strategy for preventing or reducing graft rejection in a recipient of a non-syngeneic graft which has been proposed involves administration to the recipient of CD4+CD25+ cells. Hematopoietic cells which are CD4+CD25+ have been shown to be essential for the induction and maintenance of self-tolerance and for the prevention of autoimmunity (Shevach, E. M., 2001. J Exp Med 193:F41; Shevach, E. M., 2000. Annu Rev Immunol 18:423). For example, such cells prevent the activation and proliferation of autoreactive T-cells that have escaped thymic deletion or recognize extrathymic antigens. It has been reported that CD4+CD25+ cells play a major role in tolerance induction to allogeneic responses (Graca, L., S. et al., 2002. J Exp Med 195:1641; Hara, M. et al., 2001. J Immunol 166:3789; Gregori, S. et al., 2001. J Immunol 167:1945; Chiffoleau, E. et al., 2002. J Immunol 168:5058).
Various approaches have been attempted for using CD4+CD25+ cells for inducing tolerance to non-syngeneic grafts. It has been demonstrated that donor-type CD4+CD25+ cells can be used in strategies aimed at controlling GVHD following allogeneic bone marrow transplantation in mice (Cohen, J. L. et al., 2002. J Exp Med 196:401; Hoffmann, P. et al., 2002. J Exp Med 196:389; Taylor, P. A. et al., 2002. Blood 99:3493). For example, addition of freshly isolated donor CD4+CD25+ cells to donor inoculum containing alloreactive T-cells efficiently has been shown to prevent graft-versus-host disease (GVHD) in lethally irradiated mice. The ability of CD4+CD25+ cells to promote engraftment of bone marrow allografts has been demonstrated using either host-type (Joffre, O. et al., 2004. Blood 103:4216) or donor-type CD4+CD25+ cells (Hanash, A. M., and R. B. Levy. 2005. Blood 105:1828). Induction of bone marrow allograft tolerance by co-administration to the graft recipient of the combination of donor-type CD4+CD25+ cells and rapamycin, optimally further in combination with anti-third party (veto) CTLs has also been demonstrated (Steiner et al., 2003. Blood 102, Abstract #119).
Since the number of CD4+CD25+ cells available in peripheral blood or spleen is low relative to the numbers required for therapeutic purposes, various strategies for growing these cells ex-vivo have been developed. Although, CD4+CD25+ cells exhibit low proliferative potential in-vitro upon TCR stimulation (Takahashi, T. et al., 1998. Int Immunol 10:1969), the feasibility of growing mouse or human regulatory cells has been demonstrated (Taylor, P. A. et al., 2002. Blood 99:3493; Xia, G. et al., 2004. Biol Blood Marrow Transplant 10:748; Levings, M. K. et al., 2001. J Exp Med 193:1295; Godfrey, W. R. et al., 2004. Blood 104:453; Godfrey, W. R. et al., 2005. Blood 105:750; Tang, Q. et al., 2004. J Exp Med 199:1455). This has been achieved mainly using a combination of TCR stimulation (either with an anti-TCR antibody or with allogeneic stimulator cells), costimulatory signals and high doses of IL-2. Ex-vivo-expanded CD4+CD25+ cells have been shown to retain their immunosuppressive capacities following expansion (Levings, M. K. et al., 2001. J Exp Med 193:1295). Moreover, it has been shown that differentiation of CD4+CD25+ T-regulatory cells from mouse or human CD4+CD25− T-cells can be induced under a variety of conditions (Grundstrom, S. et al., 2003. J Immunol 170:5008; Chen, W. et al., 2003. J. Exp. Med. 198:1875; Zheng, S. G. et al., 2002. J Immunol 169:4183; Curotto de Lafaille, M. A. et al., 2004. J Immunol 173:7259; Walker, M. et al., 2003. J. Clin. Invest. 112:1437), and that CD4+CD25+ cells can be induced to differentiate from both naïve and memory CD4+CD25− T-cell precursors (Walker, M. R. et al., 2005. Proc. Natl. Acad. Sci. U.S.A. 102:4103). Several studies have demonstrated that administration of donor- or host-type CD4+CD25+ cells might useful for controlling in-vivo GVHD or bone marrow allograft rejection under non-myeloablative conditions (Taylor, P. A. et al., 2002. Blood 99:3493; Hoffmann, P. et al., 2002. J Exp Med 196:389; Cohen, J. L. et al., 2002. J Exp Med 196:401; Trenado, A. et al., 2003. J Clin Invest 112:1688; Anderson, B. E. et al., 2004. Blood 104:1565; Johnson, B. D. et al., 2002. Biol Blood Marrow Transplant 8:525; Jones, S. C. et al., 2003. Biol Blood Marrow Transplant 9:243; Joffre, O. et al., 2004. Blood 103:4216; Hanash, A. M., and R. B. Levy, 2005. Blood 105:1828; Taylor, P. A. et al., 2004. Blood 104:3804).
Thus, the prior art fails to provide a satisfactory/optimal method of using tolerogenic cells, such as CD4+CD25+ cells, to prevent or reduce in a subject rejection of a therapeutic graft which is non-syngeneic with the subject.
There is thus a widely recognized need for, and it would be highly advantageous to have, a method devoid of the above limitation.